Production and Purification of Alkaline Protease from Bacillus sp. MPTK 712 Isolated from Dairy Sludge

Abstract

3 Abstract: Proteolytic bacteria were isolated from dairy sludge with the help of Skim milk agar plates. The isolate showing maximum activity was selected (MPTK 712) and characterized based on Bergey's manual. Upon 16S rDNA analysis, it displayed maximum similarity with Bacillus spp. and the sequence has been deposited in Genbank. For the production of alkaline protease, the strain MPTK 712 was grown on modified production medium containing dairy sludge as substrate. Different cultural parameters were optimized for maximal enzyme production. Peak proteolytic activity was observed at pH-9.0 and temperature at 55°C with 2% inoculums. Cetrimide exhibited the highest inhibitory activity followed by Tween-80, SDS and Tween-20. Alkaline protease was purified to homogeneity by Q-Sepharose revealing a molecular weight of 24 kDa. The enzyme was active and more stable at pH-9 and 55°C.